Current therapies for chronic HCV infections are inadequate because of low response rates, toxic side effects, and unsustained viral load reductions. As with other chronic infections (HBV and HIV-1), long-term therapy with multiple drugs will most likely be required to successfully treat chronic HCV infections and significantly reduce or eliminate progressive hepatocellular damage and hepatocellular carcinoma. The only licensed therapy for chronic HCV is interferon (IFN)-alpha, either alone or in combination with ribavirin. Combination therapy with ribavirin and IFN-alpha for 6 to 12 months is currently the treatment of choice for HCV infection. The overall sustained response rate to treatment, defined as loss of HCV from serum 6 months after completion of treatment, is 40%. Thus, there is an urgent need for better agents to treat chronic HCV infections. We have designed a novel antiviral against HCV screening technology using the HCV replicon system. Using this approach we identified modified nucleoside analogues with potent and selective in vitro anti-HCV activity. In this grant proposal, we plan to design and synthesize a total of one hundred and ninety novel 2'-C- and/or 4'-C-modified nucleosides, as well as 3'-deoxynucleosides as potential anti-HCV agents. We will determine the anti-HCV activity of a series of newly designed compounds in vitro. In addition, in preparation for in vivo proof of principle studies, adequate safety and favorable pharmacokinetic (PK) profiles of candidate compounds will be determined in relevant animal models. Furthermore, potent HCV polymerase inhibitors will be used to select for drug-resistant viral mutants, and therefore, selection of HCV replicons with the proper mutations will be a relevant part of this proposal. [unreadable] [unreadable] [unreadable]